The mood stabilizer lithium inhibits a select group of enzymes, including glycogen synthase kinase-3 (GSK-3). However, it is unclear if lithium inhibition of GSK-3 is relevant for its antimanic and antidepressant effectiveness. We are utilizing biochemical, cellular, histochemical, genomic, and behavioral validation approaches to investigate whether the inhibition of GSK-3 is an integral part of the mechanism of lithiums clinical effects. We are utilizing rodent behavioral models and two distinct but complementary approaches (pharmacologic inhibition and transgenic gene expression) in an attempt to further validate GSK-3 as a possible mediator of lithiums therapeutic effects. Specifically, one of the primary targets of GSK-3 is the transcription factor beta-catenin. We have shown that lithium administration to rats in a clinically relevant paradigm results in an increase in beta-catenin levels. We are studying the effects of both over-expression and under-expression of beta-catenin in the mouse brain as well as pharmacological mechanisms to increase beta-catenin. Using these approaches, we have found that rodents exhibit both antidepressant-like and antimanic-like behavior after treatment. Combined, these data support the hypothesis that lithium may exert its antidepressant and antimanic effects through inhibition of GSK-3, and that novel small-molecule GSK-3 inhibitors may represent a truly novel class of medications useful for the treatment of bipolar disorder and depression. Validation of lithiums therapeutic target will require clinical trials with novel inhibitors, the development of which is in progress. Like lithium, GSK-3 inhibitors exert both antimanic and antidepressant efficacy. However, the molecular mechanism through which GSK-3 inhibition influences emotional behavior is unknown. Accumulating evidence suggests that glutamatergic synaptic transmission plays a major role in the pathophysiology and treatment of bipolar disorder, suggesting that elucidation of the mechanism via which GSK-3 modulates synaptic plasticity, and particularly glutamatergic synaptic transmission, may illuminate the common pathway whereby GSK-3 inhibitors produce mood stabilizing effects. We found that GSK-3 inhibitors AR-A014418 regulated the cellular internalization of a glutamate receptor subtype known as the alpha-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate (AMPA) receptor, including Glutamate receptor 1 (GluR1) and Glutamate Receptor 2 (GluR2). We found that GluR1 and GluR2 internalization occurs via phosphorylation of kinesin light chain 2 (KLC2), the key molecule of the kinesin cargo delivery system. Specifically, AMPA stimulation triggered serine phosphorylation of KLC2 and, subsequently, the dissociation of the GluR1/KLC2 protein complex. This suggests that GSK-3 phosphorylation of KLC2 led to the dissociation of AMPA-containing vesicles from the kinesin cargo system. The peptide TAT-KLCpCDK, a specific inhibitor for KLC2 phosphorylation by GSK-3beta, reduced both the formation of a memory-related process called long-term depression (LTD) and the internalization of AMPA receptors. Furthermore, the TAT-KLCpCDK peptide showed an anti-manic effect similar to lithium on amphetamine-induced hyperactivity in mice, as well as an antidepressant effect in the tail suspension test. Taken together, the results identified a cellular mechanism, KLC2 phosphorylation, as a novel target of GSK-3beta to regulate synaptic plasticity, particularly associated with AMPA receptor trafficking and mood-associated behaviors. This kinesin cargo system provides valuable novel targets for the development of new therapeutics for mood disorders. The findings are also the first to elucidate the molecular mechanism of action for lithium's mood stabilizing effects.